Devices to test biological samples may have a sample-receiving zone that is distinct from the testing zone. As a non-limiting example, devices employing lateral flow technology have a sample-receiving zone, and the analytes flow towards the reaction zone. To retain red blood cells and/or other sample components that may interfere with the assay, a filter may be placed at or near the sample-receiving zone. However, cells or particulate materials escaping the filter can move through the device, causing high background and/or assay interference. Those limitations are more pronounced when the sample volume is variable, for example when the sample is blood obtained by pricking a finger, or when the amount of particulates in the sample is not exactly known. The sample size variability adds a need to pipette blood samples for volume consistency (Laderman, E. I. et al. 2008 Clinical and Vaccine Immunology 15: 159-163; expressly incorporated by reference herein), which is both unpractical and can result in errors during routine point of care and/or patient self-testing.
Volumes ranging from less than 1 μl to more than 20 μl can result from a finger pricking, depending on factors such as the finger (Thumb, index, middle, ring or baby finger), gender (male or female), size of the hand, blood vessels at the specific area where the lancet is introduced, and pressure applied to the finger. Those sources of variability can render use of filters to separate red blood cells ineffective: filter of one given size and thickness may be too small to retain all blood cells from a large blood drop, resulting in leakage; conversely, if the filter is made too large, it may retain samples, preventing them form reaching a test zone. The use of a pre-determined volume of blood requires blood collection, followed by pipetting of the desired volume. Besides, in some cases multiple finger pricking may be required to collect the required blood volume. Variability is also intrinsic to other complex samples; dilution with a liquid may be necessary to make the analyte available for testing. As a non-limiting example, grounded biological tissues can display uneven characteristics in terms of amount of particulates versus other components.
An effective point of care device for testing blood samples should be simple to operate, allowing either patients or health care providers to consistently execute the test by pricking a finger once, without requiring, for example, multiple finger pricking to separate plasma prior to the test. There is also an unmet need for said devices and rapid diagnostics for testing a wide variety of complex samples, containing solid or particulate matter. In addition, the device should also be effective when used with less complex samples, for example plasma or other fluids.
To avoid hook effect and/or to allow optimal flow of analytes from the sample, and/or to minimize matrix assay interference, a dilution of the sample is often times required in assays. “Hook effect”, as defined herein, is meant assay interference caused by very high concentrations of analyte. Another limitation with currently available device formats is that they do not allow for consistent flow of buffers, and hence introduce the need for pipetting in order to add measured buffers to the device. That is not practical for one unskilled in the art to perform, and also can add additional variability due to factors such as pipetting mistakes, buffer preparation, and calibration.
Although filters and complex materials such as ones used for separating erythrocytes from whole blood are known in the art [U.S. Pat. No. 5,186,843; expressly incorporated by reference herein], they do not solve the above-described limitations.
In addition, when liquid is applied directly over the sample-receiving zone, an additional problem is created, in that the liquid flow further favors movement of cells and/or other particulates towards the testing zone.
Testing of biological samples such as tissues suffer from similar limitations as described above. It should be noted that in the case of blood and/or some other complex samples, the possibility of contamination with contagious agents might render unpractical a device that needs to be assembled during use.
There is an unmet need for a simple analyte-detecting device with the following characteristics: (i) can be used by ones not skilled in the art, for example patients, or point of care providers; (ii) the parts are contained within the device, no need to pipette liquids; (iii) is operable within a range of sample volumes, thereby being suitable for samples such as one small blood drop (less than 5 μl), while still providing acceptable results with larger sample volumes.